The candidate Dr. Pingo Huang proposes highly original studies of the physiological regulation of the cystic fibrosis transmembrane conductance regulator (CFTR) by tightly compartmentalized signaling pathway in the apical membrane of human airway epithelia. The candidate is uniquely qualified due to his extensive training in protein chemistry and in patch-clamp electrophysiology. He will work at the Cystic Fibrosis Pulmonary Research Center at UNC (Director, Richard Boucher, M.D.) under the direction of M. Jackson Stutts, Ph.D. This is a strong and nurturing environment for the development of the candidate s career. He interacts on a daily basis with internationally recognized CF investigators and cell biologists such as Sharon Milgram, Ph.D. The candidate s long-term goal is to carry out independent research on apical membrane compartmentalized signal tranduction pathways that are important for CFTR function and pathophysiology of epithelial diseases. His immediate career goal is to establish an independent track record of publications and funding based on the research proposed here. The research he proposed is an exciting hybrid of cell biology and electrophysiology. This experience and the opportunities during this award period (Gordon Conferences, Carolina Workshop) will put him at the forefront on modern techniques of studying epithelial pathophysiology. The work proposed here is based on the candidate s seminal observation that the excised patch clamp technique is a highly effective approach for isolating the signal transduction mechanisms closely associated with CFTR C1- channels in the apical membrane of polarized epithelial cells. The candidate has recognized that this maneuver not only identifies signaling elements compartmentalized in the apical membrane, but also uses the activity of CFTR itself as the highly relevant physiologic readout. The main hypothesis in the proposal is that coupling of luminal adenosine receptors to CFTR in human airway epithelia is mediated by compartmentalized and apical membrane- associated PKA and PKC mediated signaling pathways. Specific aims are: 1. Determine if luminal adenosine receptors are coupled to CFTR by compartmentalized and apical membrane-associated signaling pathways. 2. Determine if a PKA singling pathway is involved in the compartmentalized and apical membrane-associated signaling from adenosine receptors to CFTR. 3. Determine if a PKC signaling pathway is involved in the compartmentalized and apical membrane-associated signaling from adenosine receptors with CFTR. The results will provide novel insight into the mechanisms of not only regulation of CFTR by adenosine in particular, but also signal transduction in the apical pole of epithelial cells in general. Furthermore, these studies could potentially provide new maneuvers for pharmacotherapy of cystic fibrosis lung disease.